Using LC/ESI-MS/MS method that was developed during the past review period, we continued to evaluate possible contribution of acute ethanol infusion and genetic background to SAL levels in human plasma and rat brain tissues. We also evaluated SAL and DA levels in plasma and CSF samples of alcoholic patients in relation to relapse behavior. We found that the levels of both SAL isomers and DA were significantly lower in the nucleus accumbens (NAC) of alcohol-preferring (P) rats in comparison to non-alcohol-preferring (NP) rats. There were decreasing trends of SAL in the striatum (STR) and DA in both NAC and STR of P rats after chronic free-choice ethanol drinking. However, the SAL levels in NAC of P rats remained unchanged. Similarly, acute ethanol infusion did not change SAL levels in plasma from healthy humans. No changes in enantiomeric ratio were observed with the acute or chronic ethanol exposure cases. These data suggest that alcohol ingestion may not change the SAL levels. Instead, lower SAL and DA levels in NAC may be associated with innate alcohol preference, influencing alcohol relapse behavior. Analysis of human plasma and CSF samples indicated that plasma and CSF SAL levels correlate very well (S-SAL, r2=0.67;R-SAL, r2=0.69), while DA levels do not. To explore whether SAL can be a possible relapse predictor in recovering alcoholics, enantiomeric (R/S)-SAL and DA levels in plasma and CSF were determined using LC/ESI-MS/MS after 3 weeks of detoxification, and drinking behavior was evaluated after 12 weeks. No statistically significant correlation was found between the SAL level in the plasma or CSF and the outcome of relapse. However, the CSF to plasma SAL ratio significantly correlated with the total days of drinking (r=-0.44, n=49;p<0.002). Our data from alcoholic patients suggested that the CSF SAL to plasma SAL ratio, as an indicator for abnormal DA metabolism in the brain, may be a possible relapse predictor in human alcoholics.